Biology subjects

Chara, O.

Publications and source records attributed to Chara, O..

17 recordsLinked to original sources

bioRxiv2026

Claudin-dependent clonal expansion couples epicardial activation to regenerative fate acquisition

Epicardial activation is a conserved response to injury across vertebrates, yet only in regenerative species do epicardium-derived cells (EPDCs) acquire the capacity to invade the injury site and contribute to cardiac regeneration. How regenerative competence is established in epicardial cells downstream of activation remains poorly understood. Using multicolour lineage tracing we find that epicardial cells of the salamander species Pleurodeles waltl undergo a transient phase of clonal expansion that precedes EPDC accumulation at the injury site. Depletion of cCpE-sensitive Claudins from otherwise intact tight junctions alters the organisation and the mechanical state of the epicardium, impairs clonal expansion and reduces the capacity of epicardial cells to delaminate. Consequently, EPDCs fail to activate a regenerative transcriptional programme and complete their transition into injury-induced migratory EPDCs that are required to sculpt a Tenascin-C-rich matrix supporting cardiomyocyte protrusion at the injury site. Together, our findings demonstrate that epicardial activation alone is insufficient for regeneration and identify Claudin-dependent clonal expansion as a mechanism that couples epicardial activation to the acquisition of a regenerative EPDC state.

cell biology↗

bioRxiv2026

StressNET: an adaptable deep-learning model for mechanical stress inference in tissues

Mechanical interactions between cells are fundamental to tissue morphogenesis during development and regeneration. Computational methods that infer intercellular stresses from microscopy images of cell shapes offer a non-invasive alternative to experimental perturbation techniques, yet all existing approaches rely on explicit physical models. Here we present StressNET, a Graph Neural Network (GNN) that infers intercellular mechanical stresses directly from tissue geometry, without assuming any underlying physical model. We generated synthetic datasets to train and benchmark StressNET, and demonstrate that its predictions achieve state-of-the-art correlation with experimental stress proxies in zebrafish neuromasts and Xenopus embryos. Analysis of the network's latent space reveals that StressNET learns global organizational principles of mechanical stress distribution, beyond local cell-cell interactions. StressNET is open-source and provides pre-trained models that can be fine-tuned on in vivo data, making it a broadly adaptable tool for studying tissue mechanics across biological systems.

bioinformatics↗

bioRxiv2026

Long-Range Coupling of Posterior Cell Addition and Anterior Vacuolation Provides Robustness in Notochord Elongation.

Robust tissue growth control requires long-range communication between the rate of progenitor addition and tissue expansion. However, the regulatory mechanisms that couple these two processes are unknown. In zebrafish, notochord morphogenesis is a principal driver of axis extension through the combined actions of posterior progenitor addition and anterior vacuolation. To elucidate how progenitor dynamics and vacuole-driven cell expansion interact to shape notochord development, we first generated a mathematical model that links progenitor addition rate to the progressive expansion of cells from anterior-to-posterior to simulate vacuolation rate. Comparing this with empirical measurements, we find that progenitor incorporation together with vacuolation, produces a linear gradient in nearest neighbour distance. We next explored the role of YAP/TAZ in regulating the rate of progenitor addition in mutants for YAP/TAZ inhibitor vgll4b. We find that vgll4b expression and YAP activity are enriched in posterior midline progenitors. Loss of vgll4b elevates YAP signaling, enhances progenitor addition, restricts vacuole expansion, and--after a transient buffering phase--compromises A-P axis elongation. These results support a long-range feedback mechanism linking progenitor recruitment to vacuolation, enabling the notochord to balance cellular input with volumetric expansion, thereby maintaining tissue proportions. Graphical abstract. Proposed working model O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=166 SRC="FIGDIR/small/706348v2_ufig1.gif" ALT="Figure 1"> View larger version (62K): org.highwire.dtl.DTLVardef@6c3c6dorg.highwire.dtl.DTLVardef@1f34eb0org.highwire.dtl.DTLVardef@b33a21org.highwire.dtl.DTLVardef@ad582b_HPS_FORMAT_FIGEXP M_FIG C_FIG A. Schematic representation of the authors proposed model illustrating how increased YAP activation in notochord progenitors of vgll4b mutants leads to enhanced progenitor addition to the notochord. This increased incorporation subsequently compromises the ability of notochord cells to undergo proper vacuolation, resulting in reduced axial elongation compared with control embryos.

developmental biology↗

bioRxiv2025

Brachyury expression levels predict lineage potential and axis-forming ability of in vitro derived neuromesodermal progenitors

Neuromesodermal progenitors (NMPs) produce the spinal cord and musculoskeleton in the elongating anterior-posterior axis. In vivo, NMPs possess dual potency, coinciding with regions coexpressing SOX2 and Brachyury (TBXT). In vitro, SOX2/TBXT co-expressing cells can be produced from pluripotent cells and, like their in vivo counterparts, can produce neural tube and somitic mesoderm. However, the functional characteristics of in vitro SOX2/TBXT co-expressing cells remain unclear, confounding comparisons with in vivo data. To address this, we developed a dual Sox2/Tbxt reporter mouse ESC line. SOX2/TBXT reporter-positive cells emerge in vitro from pluripotent populations with dynamics that mirror their appearance in the embryo. Purified SOX2/TBXT co-expressing populations can differentiate towards neurectoderm or mesoderm, including lateral mesoderm upon BMP stimulation. In gastruloids, quantitative live imaging shows that WNT or NOTCH inhibition rapidly leads to downregulation of TBXT expression and diminished axial extension. We show that clonally plated SOX2/TBXT co-expressing cells are bipotent NMPs that can also self-propagate. By combining clonal analysis with mathematical modelling, we identify two thresholds of SOX2/TBXT expression, switching clonal output from neural- to mesoderm-biased, and from mesoderm-biased to mesoderm-specified. Media and substrate composition alter the lineage outcomes of in vitro derived mouse NMPs. Thus, this Sox2/Tbxt double reporter cell line provides support for unsuspected heterogeneity in NMPs, together with evidence for a role of these transcription factors in directing cell fate to drive axis elongation.

developmental biology↗

bioRxiv2025

Mechanical control of tissue growth during limb regeneration

The axolotl is a highly regenerative species, capable of restoring full limbs, regardless of the amputation site. However, the regeneration rate is adjusted with the plane of amputation along the proximo-distal (PD) axis, leading to equivalent regeneration times regardless of the extent of tissue removal. We hypothesized that this phenomenon could be partly explained by differences in tissue mechanical properties. In this work, we describe tissue growth mathematically and evaluate cell cycle parameters of regenerating limbs amputated at different levels along the PD axis, demonstrating a linear correlation between the cell cycle length and the amputation site during early regeneration phases. We show as well, that blastema cells require their endogenous context to retain such proliferation differences. We measured mechanical properties in regenerating limbs with in vivo optical and standard indentation-based techniques and demonstrated that distal blastema cells are stiffer than proximal ones. Accordingly, we demonstrated that axolotl cells decrease their proliferation with increased extracellular matrix stiffness in vitro. Next, we evaluated the activity of the mechanotransducers YAP/TAZ in vivo by using a GTIIC-based reporter line combined with target gene expression data, which indicated that their activity peaks during the blastema stage, with higher activity after proximal amputations. Hence, our findings strongly suggest a mechanical dependence for the position-dependent regulation of cell proliferation during axolotl limb regeneration, where YAP/TAZ likely plays a role in the mechanotransduction mechanism.

biophysics↗

bioRxiv2025

Ethylene modulates cell wall mechanics for root responses to compaction

Soil stresses impact crop yields, presenting global agricultural challenges. Soil compaction triggers root length reduction and radial expansion driven by the plant hormone ethylene. We report how ethylene controls cell wall properties to promote root radial expansion. We demonstrate how soil compaction stress, via ethylene, upregulates Auxin Response Factor1 in the root cortex, which represses Cellulose Synthase (CESA) genes. CESA repression drives radial expansion of root cortical cells by modifying the thickness and mechanics of their cell walls, which result in a "stiff epidermis-soft cortex" contrast. Our research thus connects ethylene signaling with root mechanics via cell wall strength, and reveals how dynamic regulation of cellulose synthesis crucially controls root growth in compacted soil.

plant biology↗

bioRxiv2024

Local control of cellular proliferation underlies neuromast regeneration in zebrafish

Biological systems are never in equilibrium but maintain stability despite continuous external disturbances. A prime example of this is organ regeneration, where, despite intrinsically stochastic damage, organs are rebuilt through controlled cellular proliferation. In this study, we employ a cell- based computational modelling approach to investigate the proliferative response to injury. We developed a minimal two-dimensional Cellular Potts Model (CPM) using empirical data from regenerating neuromasts in larval zebrafish. Remarkably, the CPM both qualitatively and quantitatively recapitulates the regenerative response of neuromasts following laser-mediated cell ablation. Assuming that cell proliferation is locally regulated by a delayed switch, mitotic activity ceases once the type-dependent number of neighbouring cells exceeds a deterministic critical threshold. An intriguing corollary of our findings is that a local negative feedback loop among identical cells may represent a general mechanism underlying organ-level proportional homeostasis.

developmental biology↗

bioRxiv2024

Lineage tracing of Shh+ floor plate cells and dynamics of dorsal-ventral gene expression in the regenerating axolotl spinal cord

Both development and regeneration depend on signalling centres, which are sources of locally secreted tissue-patterning molecules. As many signalling centres are decommissioned before the end of embryogenesis, a fundamental question is how signalling centres can be re-induced later in life to promote regeneration after injury. Here, we use the axolotl salamander model (Ambystoma mexicanum) to address how the floor plate is assembled for spinal cord regeneration. The floor plate is an archetypal vertebrate signalling centre that secretes Shh ligand and patterns neural progenitor cells during embryogenesis. Unlike mammals, axolotls continue to express floor plate genes (including Shh) and downstream dorsal-ventral patterning genes in their spinal cord throughout life, including at steady state. The parsimonious hypothesis that Shh+ cells give rise to functional floor plate cells for regeneration had not been tested. Using HCR in situ hybridisation and mathematical modelling, we first quantitated the behaviours of dorsal-ventral spinal cord domains, identifying significant increases in gene expression level and floor plate size during regeneration. Next, we established a transgenic axolotl to specifically label and fate map Shh+ cells in vivo. We found that labelled Shh+ cells gave rise to regeneration floor plate, and not to other neural progenitor domains, after tail amputation. Thus, despite changes in domain size and downstream patterning gene expression, Shh+ cells retain their floor plate identity during regeneration, acting as a stable cellular source for this regeneration signalling centre in the axolotl spinal cord.

developmental biology↗

bioRxiv2024

ForSys: non-invasive stress inference from time-lapse microscopy

During tissue development and regeneration, cells interpret and exert mechanical forces that are challenging to measure in vivo. Therefore, stress inference algorithms have emerged as powerful tools to estimate tissue stresses. However, how to incorporate tissue dynamics effectively into the inference remains elusive. Here, we present ForSys, a Python-based software that estimates intercellular stresses and intracellular pressures using time-lapse microscopy. We validated ForSys in silico and in vivo using the well-characterized mucociliary epithelium of the Xenopus embryo. We applied ForSys to study the migrating zebrafish lateral line primordium. We found that stress increases during cell rounding just before cell division and predicted the onset of epithelial rosettogenesis with high accuracy. Finally, we analyzed the development of the zebrafish neuromast and inferred mechanical asymmetries in a cell type-specific adhesion pattern. The versatility and simplicity of ForSys enhance the toolkit for studying spatiotemporal patterns of mechanical forces during tissue morphogenesis in vivo.

systems biology↗

bioRxiv2023

How a reaction-diffusion signal can control spinal cord regeneration in axolotls: A modelling study

Axolotls are uniquely able to completely regenerate the spinal cord after amputation. The underlying governing mechanisms of this regenerative response have not yet been fully elucidated. We previously found that spinal cord regeneration is mainly driven by cell cycle acceleration of ependymal cells, recruited by a hypothetical signal propagating from the injury. However, the nature of the signal and its propagation remain unknown. In this theoretical study, we investigated whether the regeneration-inducing signal can follow a reaction-diffusion process. We developed a computational model, validated it with experimental data and showed that the signal dynamics can be understood in terms of reaction-diffusion mechanism. By developing a theory of the regenerating outgrowth in the limit of fast reaction-diffusion, we demonstrate that control of regenerative response solely relies on cell-to-signal sensitivity and the signal reaction-diffusion characteristic length. This study lays foundations for further identification of the signal controlling regeneration of the spinal cord.

developmental biology↗

bioRxiv2023

Fluctuations in tissue growth portray homeostasis as a critical state and long-time non-Markovian cell proliferation as Markovian

Tissue growth is an emerging phenomenon that results from the cell-level interplay between proliferation and apoptosis, which is crucial during embryonic development, tissue regeneration, as well as in pathological conditions such as cancer. In this theoretical article, we address the problem of stochasticity in tissue growth by first considering a minimal Markovian model of tissue size, quantified as the number of cells in a simulated tissue, subjected to both proliferation and apoptosis. We find two dynamic phases, growth and decay, separated by a critical state representing a homeostatic tissue. Since the main limitation of the Markovian model is its neglect of the cell cycle, we incorporated a refractory period that temporarily prevents proliferation immediately following cell division, as a minimal proxy for the cell cycle, and studied the model in the growth phase. Importantly, we obtained from this last model an effective Markovian rate, which accurately describes general trends of tissue size. This study shows that the dynamics of tissue growth can be theoretically conceptualized as a Markovian process where homeostasis is a critical state flanked by decay and growth phases. Notably, in the growing non-Markovian model, a Markovian-like growth process emerges at large time scales.

biophysics↗

bioRxiv2022

Apical-to-basal graded ROS metabolism in intact Hydra leads to distinct levels of injury-induced ROS signaling in apical and basal regenerating tips

After mid-gastric bisection, Hydra regenerates a head from the lower half and a basal disc from the upper one. What signals elicit two distinct regenerative responses in bisected Hydra remains unknown. A mathematical modeling approach based on quantitative data linked to MAPK activation and injury-induced cell death predicts an immediate release of a locally restricted short-lived signal in apical-regenerating tips. We found that Reactive oxygen species (ROS) fulfill this role as evidenced by the injury-induced production of hydrogen peroxide (H2O2), three-fold higher in apical-regenerating tips than in basal ones. By contrast, mitochondrial superoxide (mtO2.-) is similarly produced on each side of the cut, playing a positive role on wound healing as mtO2.- scavenging delays healing whereas knocking-down Super Oxide Dismutase (SOD) leads to mtO2.- accumulation and acceleration of wound-healing. In intact Hydra, the ROS-processing enzyme activities are inversely graded along the body column, basal-to-apical for SOD and apical-to-basal for catalase, explaining the asymmetrical levels of H2O2 after bisection. High H2O2 levels trigger injury-induced cell death via paracrine signaling in apical-regenerating tips, where NOX4 and CYBB enzymes amplify them. Hence, the asymmetrical regulation of H2O2 levels immediately after amputation is crucial to activate two distinct regenerative responses in Hydra.

developmental biology↗

bioRxiv2021

Size matters: an analytical study on the role of tissue size in spatiotemporal distribution of morphogens unveils a transition between different reaction-diffusion regimes

The reaction-diffusion model constitutes one of the most influential mathematical models to study distribution of morphogens in tissues. Despite its widespread use, the effect of finite tissue size on model-predicted spatiotemporal morphogen distributions has not been completely elucidated. In this study, we analytically investigated the spatiotemporal distributions of morphogens predicted by a reaction-diffusion model in a finite 1D domain, as a proxy for a biological tissue, and compared it with the solution of the infinite-domain model. We explored the reduced parameter, the tissue length in units of a characteristic reaction-diffusion length, and identified two reaction-diffusion regimes separated by a crossover tissue size estimated in [~]3.3 characteristic reaction-diffusion lengths. While above this crossover the infinite-domain model constitutes a good approximation, it breaks below this crossover, whereas the finite-domain model faithfully describes the entire parameter space. We evaluated whether the infinite-domain model renders accurate estimations of diffusion coefficients when fitted to finite spatial profiles, a procedure typically followed in Fluorescence Recovery After Photobleaching (FRAP) experiments. We found that the infinite-domain model overestimates diffusion coefficients when the domain is smaller than the crossover tissue size. Thus, the crossover tissue size may be instrumental in selecting the suitable reaction-diffusion model to study tissue morphogenesis.

systems biology↗

bioRxiv2021

Tig1 regulates proximo-distal identity during salamander limb regeneration

Salamander limb regeneration is an accurate process which gives rise exclusively to the missing structures, irrespective of the amputation level. This suggests that cells in the stump have an awareness of their spatial location, a property termed positional identity. Little is known about how positional identity is encoded, in salamanders or other biological systems. Through single-cell RNAseq analysis, we identified Tig1/Rarres1 as a potential determinant of proximal identity. Tig1 encodes a conserved cell surface molecule, is regulated by retinoic acid and exhibits a graded expression along the proximo-distal axis of the limb. Its overexpression leads to regeneration defects in the distal elements and elicits proximal displacement of blastema cells, while its neutralisation blocks proximo-distal cell surface interactions. Critically, Tig1 reprogrammes distal cells to a proximal identity, upregulating Prod1 and inhibiting Hoxa13 and distal transcriptional networks. Thus, Tig1 is a central cell surface determinant of proximal identity in the salamander limb.

developmental biology↗

bioRxiv2020

Modeling the spatiotemporal control of cell cycle acceleration during axolotl spinal cord regeneration

Axolotls are uniquely able to resolve spinal cord injuries, but little is known about the mechanisms underlying spinal cord regeneration. We previously found that tail amputation leads to reactivation of a developmental-like program in spinal cord ependymal cells (Rodrigo Albors et al., 2015), characterized by a high-proliferation zone emerging 4 days post-amputation (Rost et al., 2016). What underlies this spatiotemporal pattern of cell proliferation, however, remained unknown. Here, we use modelling, tightly linked to experimental data, to demonstrate that this regenerative response is consistent with a signal that recruits ependymal cells during 85 hours after amputation within ~830{square}m of the injury. We adapted FUCCI technology to axolotls (AxFUCCI) to visualize cell cycles in vivo. AxFUCCI axolotls confirmed the predicted appearance time and size of the injury-induced recruitment zone and revealed cell cycle synchrony between ependymal cells. Our modeling and imaging move us closer to understanding bona fide spinal cord regeneration.

bioinformatics↗

bioRxiv2019

A dynamic cell recruitment process drives growth of the Drosophila wing by overscaling the Vestigial expression pattern

Organs mainly attain their size by cell growth and proliferation, but sometimes also grow through recruitment of undifferentiated cells. Here we investigate the participation of cell recruitment in establishing the pattern of Vestigial (Vg), the product of the wing selector gene in Drosophila. We find that the Vg pattern overscales along the dorsal-ventral (DV) axis of the wing imaginal disc, i.e., it expands faster than the DV length of the pouch. The overscaling of the Vg pattern cannot be explained by differential proliferation, apoptosis, or oriented-cell divisions, but can be recapitulated by a mathematical model that explicitly considers cell recruitment. By impairing cell recruitment genetically, we find that the Vg pattern almost perfectly scales and adult wings are approximately 20% smaller. Furthermore, using fluorescent reporter tools, we provide direct evidence that cell recruitment takes place in a specific time between early and mid third-instar larval development. Altogether, our work quantitatively shows when, how, and by how much cell recruitment shapes the Vg pattern and drives growth of the Drosophila wing.

developmental biology↗

bioRxiv2017

Signaling-dependent control of apical membrane size and self-renewal in rosette-stage human neuroepithelial stem cells

In the early developing nervous system, self-renewing neural stem cells are polarized and maintain an apical domain facing a central lumen. The presence of apical membrane is thought to have a profound influence on maintaining the stem cell state. With the onset of neurogenesis cells lose their polarization and the concomitant loss of the apical domain coincides with a loss of the stem cell identity. Very little is known about the molecular signals controlling apical membrane size. Here we use two neuroepithelial cell systems, one derived from regenerating axolotl spinal cord and the other from human ESCs to identify a conserved molecular signalling pathway initiated by lysophosphatidic acid (LPA) that controls apical membrane size and consequently controls and maintains epithelial organization and lumen size in neuroepithelial rosettes. This apical domain size increase occurs independently of effects on proliferation and involves a SRF-dependent transcriptional induction of junctional and apical membrane components.

neuroscience↗